Many hormonal responses are mediated through c-AMP, and the major effect of c-AMP in eukaryotic cells is its interaction with c-AMP- dependent protein kinase. In the absence of c-AMP the enzyme exists as an inactive complex containing both regulatory (R) and catalytic (C) subunits. Cyclic-AMP binds to the R-subunit causing the molecule to dissociate into free C and free R subunits. The free catalytic subunit is now fully active and in the presence of ATP phosphorylates a variety of substrates including histones, caseine, protamine, phosphorylase b kinase, glycogen synthase I, and lipase. The effect is to promote an increase in glycogenolysis, lipolysis or protein synthesis. Our ultimate objective is to determine the complete amino-acid sequence of both the catalytic and regulatory subunits of c-AMP- dependent protein kinase purified from pig muscle. The first goal has been to purify the enzyme on a large scale. Subsequently the exact nature of the subunit structure will be established. The smallest component is the catalytic subunit with a molecular weight of 35,000 daltons. Additional studies including SDS gel electrophoresis, ultracentrifugation in the presence of urea, and C-terminal and N- terminal amino acid identification will establish the exact size and number of polypeptide chains in each subunit. Peptide maps of the two subunits will be characterized in detail both as a supplementary means of determining the minimum molecular weight components and as a future reference for comparative studies with other protein kinases and c-AMP receptor proteins. In conjunction with sequencing, active-site labelling experiments will be undertaken in an attempt to identify specific amino acid residues which may be essential for enzymatic activity, c-AMP binding, or subunit interaction. Extensive efforts also will be made to crystallize the enzyme. This will be essential since only when the primary structure and the crystal structure of a molecule are known, can the tertiary structure be fully appreciated. The X-ray studies will be carried out in the crystallographic of our Department at U.C.S.D.